Microrna-16 Regulates BMI1 in the Clonogenic Side Population of Refractory Mantle Cell Lymphoma

Mantle cell lymphoma (MCL) is categorized as an indolent CD5⁺ B-cell lymphoma and is associated with numerous genomic copy number alterations, including 9p21 deletion (CDKN2A) and 10p12 amplification (BMI1). The target gene of the 10p12 amplification has been identified as BMI1, whose overexpression is frequently observed in the blastoid variant of MCL. CDKN2A is also well-known target of BMI1 in solid tumor. So, it has been hypothesized that BMI1 regulates CDKN2A in MCL. However there are the MCL cases with both 10p12 amplification and 9p21 homozygous deletion, suggesting that BMI1 might regulate the other target gene(s). The proto-oncogene, BMI1 is crucially involved in cancer stem cell maintenance and the upregulation has been demonstrated in aggressive or relapsed cases of solid tumors. Cancer stem cells are often identified in the side population (SP) of cancer cells, which is detected based on the cell's ability to export Hoechst 33342 dye via an ATP-binding cassette (ABC) membrane transporter, which gives the SP a distinct low-staining pattern.

The aim of this study is to determine the role of BMI1 in MCL initiating cells, especially in the relapsed cases. In this presentation, we show that the SP fraction has stem cell-like characteristics and high tumorigenic potential, and that BMI1 expression is upregulated in the SP in both relapsed MCL cases and MCL cell lines. Further we show that miR-16 is upstream regulator of the BMI1 in MCL.

To determine the role of BMI1 in the pathogenesis of MCL-initiating cells, we firstly examined BMI1 expression at primary MCL cases and found that its expression is stronger in cases of recurrent MCL than at initial diagnosis. We next characterized the MCL SP and found that the SP cells exhibit cancer stem cell-like features and upregulated BMI1 expression, which appears to enhance anti-apoptosis activity. Knocking down of BMI1 increases apoptosis and reduces tumorigenicity in CDKN2A^−/− MCL cell lines (REC1 and Z138c). Subcutaneous inoculation of NOD/Shi-scid IL-2γnul (NOG) mice with CDKN2A^−/− MCL cell lines, siBMI1-expressing cells were significantly smaller than those in mice receiving control siRNA in vivo. Chip assay showed that BMI1 interacts with BCL2L11/Bim and PMAIP3/Noxa, which were recently shown to be Bmi-1 target. These results suggest that BMI1/Bmi-1 may regulate Bim and/or Noxa to inhibit apoptosis in MCL cells. Furthermore, upon screening for upstream regulator of BMI1, we found that expression of a non-cording regulatory RNA, microRNA-16 (miR-16) is weaker in MCL SP cells than in non-SP cells. To investigate relationship between BMI1 and miR-16, we transfected miR-16 into MCL cell lines, and found that it directly downregulated BMI1, leading to reductions in tumor size following in vivo lymphoma xenograft (NOG mice). Finally, we find that bortezomib, which is known to be a proteasome inhibitor, led to dose-and time- dependent reductions in Bmi-1 expression with re-upregulation of miR-16 in both cell lines and a primary sample.

We conclude that dysregulation of miR-16 and BMI1 plays a key role in lymphomagenesis by reducing MCL cell apoptosis, especially in refractory/recurrent cases via enhancement of anti apoptotic function.

No relevant conflicts of interest to declare.